Rhodopsin¨Ctransducin heteropentamer: Three-dimensional structure and biochemical characterization

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• 作者：Beata  ; Jastrzebska^a ;   ; ^{bxj27@case.edu} ; Philippe  ; Ringler^b ; David T.  ; Lodowski^a ; Vera  ; Moiseenkova-Bell^a ; Marcin  ; Golczak^a ; Shirley A.  ; Mü ; ller^b ; Krzysztof  ; Palczewski^a ;   ; ^{kxp65@case.edu} ; Andreas  ; Engel^a ;   ; ^b ;   ; ^{andreas.engel@case.edu}
• 关键词：G protein-coupled receptor ; Heterotrimeric G protein ; Photoactivated rhodopsin ; Scanning transmission electron microscopy ; Transducin
• 刊名：Journal of Structural Biology
• 出版年：2011
• 出版时间：December 2011
• 年：2011
• 卷：176
• 期：3
• 页码：387-394
• 全文大小：1265 K

文摘

The process of vision is initiated when the G protein-coupled receptor, rhodopsin (Rho), absorbs a photon and transitions to its activated Rho^{?/sup> form. Rho?/sup> binds the heterotrimeric G protein, transducin (G_t) inducing GDP to GTP exchange and G_t dissociation. Using nucleotide depletion and affinity chromatography, we trapped and purified the resulting nucleotide-free Rho?/sup>¡¤G_t complex. Quantitative SDS¨CPAGE suggested a 2:1 molar ratio of Rho?/sup> to G_t in the complex and its mass determined by scanning transmission electron microscopy was 221 ¡À 12 kDa. A 21.6 ? structure was calculated from projections of negatively stained Rho?/sup>¡¤G_t complexes. The molecular envelope thus determined accommodated two Rho molecules together with one G_t heterotrimer, corroborating the heteropentameric structure of the Rho?/sup>¡¤G_t complex.}